A-ketoglutarate oxidase

Their acute toxicity to mammals is low, but the two active substances may cause dermatitis. In rats fed daily on a diet containing quinomethionate a high cumulative toxicity was observed. A dietary level of 500 mg/kg for 90 days reduced body-weight, caused hypertrophy of the liver, and inhibited acetoacetate synthesis and the microsomal enzymes. It primarily inhibited the function of the HS-enzymes (pyruvate dehydrogenase, succinate dehydrogenase, malate dehydrogenase and a-ketoglutarate oxidase) (Carlson and DuBois, 1970). 

Study of the mechanism of the oxidation of pyruvic acid by certain bacteria led to the discovery of lipoic acid (thioctic acid) as a nutrient metabolite essential for the oxidative decarboxylation of a-keto acids. It subsequently was determined that the acetate-replacing factor " for lactic acid bacteria and protogen,a growth factor for the protozoan, Tetrahymena gelii, were also identical with lipoic acid. The occurrence of considerable quantities of lipoic acid in mammalian preparations of pyruvate and a-ketoglutarate oxidases suggests that it has the same function in animal tissues as in microorganisms. ... 

Sanadi and Littlefield have reported on a soluble a-ketoglutarate oxidase system which is likewise CoA-dependent. This system has previously been referred to, since it was used as an assay for CoA. In the presence of the oxidase, a supernatant factor, DPN, and CoA, a-ketoglutarate is oxidized to succinate and CO2. The reaction is visualized as follows ... 

Alcohol dehydrogenase (5) and leucine a-ketoglutarate transaminase (33,34) contribute to the development of aroma during black tea manufacturing. Polyphenol oxidase and peroxidase are essential to the formation of polyphenols unique to fermented teas. 

The a-ketoacid-dependent enzymes are distinguished from other non-haem iron enzymes by their absolute requirement for an a-ketoacid cofactor as well as Fe(II) and O2 for activity. They catalyse two types of reaction (Table 2.3), hydroxyla-tion and oxidation. In both, the a-ketoglutarate is decarboxylated and one oxygen atom introduced into the succinate formed in the hydroxylases, the other oxygen atom is introduced into the substrate, while in the oxidases it is found in water, together with the cyclized product. In general these enzymes require one equivalent of Fe(II) an a-ketoacid, usually a-ketoglutarate and ascorbate. Examples of these enzymes include proline 4-hydroxylase, prolyl and lysyl hydroxylase, which... 

In fact, the a-ketoglutarate/glutamate dehydrogenase is a generally applicable method for the regeneration of NAD and NADP in laboratory scale productions. Both components involved are inexpensive and stable. Quite recently, a method for the oxidation of the reduced nicotinamide coenzymes based on bacterial NAD(P)H oxidase has been described [225], This enzyme oxidizes NADH as well as NADPH with low Km values. The product of this reaction is peroxide, which tends to deactivate enzymes, but it can be destroyed simultaneously by addition of catalase. The irreversible peroxide/catalase reaction favours the ADH catalyzed oxidation reaction, and complete conversions of this reaction type are described. 

Less wide-spread than the enzymatic determination of these amino acids is the application of enzyme sensor systems for precursors or other amino acids. Collins et al. [116] described a flow injection system for the determination of a-ketoglutarate during an industrial fermentation. The system was based on glutamate dehydrogenase (Eq. (11.5)) and glutamate oxidase (Eq. (11.4))... 

The resonance Raman spectra of Fe(IV) complexes [(L)FeO]2+, where L = pentadentate pyridylamine ligands, include vFe=0 at 752 cm-1.298 Transient resonance Raman spectra of oxygen intermediates in the non-haem iron enzyme taurine/a-ketoglutarate dioxygenase, include characteristic bands at 812 (160)/787 (lsO) cm-1 and 583 (160)/555 (lsO) cm-1.299 Oxidised intermediates of cytochrome c oxidase give resonance Raman bands for vFe=0 showing that there are three different forms present.300... 

Fig. 5. Changes of liver enzyme activities after tyrosine dosage of guinea pigs tyrosine-a-ketoglutarate transaminase (- -o- -), p-hydroxyphenylpyruvate oxidase (— x—). Each point is the average of homogenates from two animals. From Knox and Goswami (Kll).

The most useful, and thus far successful, examples have involved irreversible reactions of nucleophilic functions of an enzyme s reactive site with an enzymatically activated Kcat inhibitor of a Michael-type addition reaction. The activation invariably requires participation of the enzyme s prosthetic group (e.g., flavin of monoamine oxidase) or coenzymes such as pyridoxal (vitamin B) as its phosphate, which is associated with several enzymes (e.g., threonine dehydrase, ornithine decarboxylase, a-ketoglutarate transaminase). 

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